Aptamers are single stranded DNA or RNA molecules that have emerged as a new class of bio-receptors that are analogous to antibodies in terms of binding to their target with high affinity and specificity. Aptamer sequence is capable of forming a unique 3D structure owing to the presence of the intramolecular attractions between the nucleotides such as van der waals, hydrogen bonding and hydrophobic interactions (Ku T. H. 2015). Aptamers have been identified for several analytes such as proteins, viruses, toxins, drugs, hormones, bacteria, metal ions, peptides, cells, ions and even tissues with high affinity that can reach picomolar dissociation constant (Kd). They have shown several advantages over antibodies which make them better candidates for many affinity-based applications, particularly in the development of biosensors (Song, S. et. al. 2008).
Aptamers are selected by an in vitro process which does not require the use of experimental animals, thus, it is possible to acquire aptamers for both toxic and non-immunogenic molecules. Indefinite number of aptamers can be made easily and at low cost once the aptamer for the target is identified and its sequence is obtained. Aptamers have high thermal stability and can be effortlessly modified by numerous chemical tags with negligible effect on their binding affinity and specificity allowing them to be easily immobilized on different solid supports. However, despite the advantages and the great promise of aptamers in different therapeutic and diagnostic applications, yet limited number of aptamers has been successfully identified, particularly for small molecules (McKeague, M et. al. 2012, Ruscito, A et. al. 2016). This is majorly attributed to the limitation of the conventional selection process of the aptamers known as Systematic Evolution of Ligands by Exponential enrichment (SELEX). The SELEX process usually involves two major steps: multiple iterative rounds of selection (10-20) from a pool of large library of DNA sequences and then amplification of the selected subpool. In each round the DNA library is incubated with the target immobilized on solid matrix (usually sugar based or magnetic microbeads). Then, several washing steps for the unbound DNA is performed followed by elution, PCR amplification of the bound DNA and then purification of the single stranded DNA pool for further rounds. Moreover, the evaluation of the aptamer enrichment is usually done by monitoring the fluorescence of the eluted DNA after each round. Thus, the SELEX process is expensive, labor intensive, time consuming and sometime inefficient.
In order to overcome these limitations, some innovative selection methods have been developed over the last 20 years combining various techniques to accelerate and improve aptamers selection (Gopinath 2007; Blind and Blank 2015). Capillary electrophoresis (CE)-SELEX (Mendosa S D et. al. 2004, Berezovski M et. al. 2005) and nitrocellulose filter binding SELEX (Blind, M et. al. 2015) have been reported. However, these methods can be only used for large molecules such as proteins. Flow cytometry-SELEX and fluorescence-activated cell sorting-SELEX (Raddatz, Dolf et al. 2008; Wang, Gong et al. 2014) have been also developed for selecting aptamers against cells. Microfluidic technologies (Hybarger, Bynum et al. 2006; Jing and Bowser 2011; Weng, Huang et al. 2012; Wang, Liu et al. 2014; Hung, Wang et al. 2016) have been also integrated with different SELEX methods such as CE (Berezovski, Drabovich et al. 2005), sol-gel and magnetic beads (Lou, Qian et al. 2009; Oh, Ahmad et al. 2011; Lai, Wang et al. 2014) in order to increase the separation efficiency. However, in these methods, the aptamer enrichment is not monitored in real-time leading to selection blindness, longer selection time and higher number of failure selection trials (Hong, Wan et al. 2017; Liu, Li et al. 2017). Therefore, it is highly demanded to develop a rapid and low cost selection approach that possesses real-time evaluation capability and remarkable efficiency.